The World Health Organization (WHO) declared a tuberculosis (TB) global emergency, and it is estimated that today 1/3 of the world population, or approximately 2 billion people, is infected with latent TB. The most dangerous form of Mycobacterium tuberculosis (Mtb) infection is central nervous system (CNS) tuberculosis (CNSTB). Despite its public health importance, current understanding about the pathogenesis of CNSTB is very limited, and prognosis for patients with CNSTB is bleak. CNSTB largely affects children and immunodeficient adults. We have established novel MHC class II- deficient or RAG immunodeficient mouse models to study Mtb dissemination in the CNS. We hypothesize that dendritic cells (DCs) deliver Mtb into the CNS and contribute to Mtb dissemination in CNSTB. In our preliminary data, we have shown that DCs preferentially carry Mtb into the CNS in 7-day-old immunodeficient mice. We have generated novel recombinant Mtb strains that express green fluorescent protein (GFP) and created Mtb strains that express OVA257-264 and OVA323-339 antigenic epitopes. We have shown that these novel Mtb strains can be tracked in vivo and utilized for testing systemic adaptive immunity in CNSTB. Our overall goal is to define the mechanisms of Mtb dissemination into the CNS. In Aim 1, we will determine the role of DCs in Mtb dissemination into the CNS (Aim 1A) and investigate the means by which the blood- brain barrier (BBB) regulates Mtb-infected DC migration (Aim 1B). In Aim 2, we will study immunological control of CNSTB. We will take advantage of our knowledge of mycobacterial control in peripheral tissues and define which CNS and immune cells control mycobacterial growth in CNSTB (Aim 2A). Using defined mouse lines transgenic for Ag85, OT-I, or OT-II, as well as recombinant Mtb strains expressing these epitopes, we will dissect the mechanism of antigen-specific immune response in CNSTB (Aim 2B). At the conclusion of these studies, we will have increased our knowledge of CNSTB pathogenesis, evaluated new in vivo models of CNSTB, created novel reagents that can be used to track Mtb infections in vivo, and elucidated the role of the BBB in Mtb dissemination into the CNS. These experiments will fill the gap in our knowledge regarding the pathogenesis of CNSTB and will lead to improved strategies for treating mycobacterial infections of the CNS. PUBLIC HEALTH RELEVANCE: Despite the availability of effective short-course chemotherapy and the Bacille Calmette- Guerin (BCG) vaccine, the tubercle bacillus continues to claim more lives than any other single infectious agent. Recent years have demonstrated an increased incidence of tuberculosis in both developing and industrialized countries, the widespread emergence of drug-resistant strains and a deadly synergy with the human immunodeficiency virus (HIV). IN 1993, the World Health Organization (WHO) declared a tuberculosis (TB) global emergency and it is estimated that today 1/3, approximately 2 billion people, of the global population is infected with latent TB. CNS tuberculosis (CNSTB) is the most dangerous form of Mtb infection. The diagnosis is challenging, and the disease is clinically hard to manage. Mortality, even with rigorous treatment, is high, and of those who survive ~27% develop severe neurological damage and disability. Despite the severity of this disease, current understanding about the pathogenesis of CNSTB is still dated back to necropsy data published in 1933. Treatment has advanced little since the introduction of isoniazid in the 1950s (Clin Chest Med 30(4): 745-754). Therapeutic approaches are limited to chemotherapy developed over 30 years ago, and the lack of progress in research on pathogenesis has put the development of new treatments on hold for decades. With drug-resistant strains emerging there is an urgent need for development of new strategies for therapeutic intervention and prevention of this disease. This project will provide new knowledge regarding pathogenesis of CNS tuberculosis and enable the development of preventive and therapeutic strategies to cope with this disease.